HaloLink™ Protein Array System

Protein arrays enable parallel analysis of multiple protein:protein, protein:drug or protein:nucleic acid interactions. The HaloLink™ Protein Array System provides a way to create homebrew (on-demand) protein arrays by combining innovative HaloTag® technology, surface engineering and cell-free protein expression systems.

The HaloTag® protein is a mutated hydrolase that forms a covalent bond with HaloTag® ligands. Under physiological conditions binding is rapid and highly specific, yielding a complex that is stable even under stringent conditions. Using the HaloLink™ Protein Array System, ...

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Protein arrays enable parallel analysis of multiple protein:protein, protein:drug or protein:nucleic acid interactions. The HaloLink™ Protein Array System provides a way to create homebrew (on-demand) protein arrays by combining innovative HaloTag® technology, surface engineering and cell-free protein expression systems.

The HaloTag® protein is a mutated hydrolase that forms a covalent bond with HaloTag® ligands. Under physiological conditions binding is rapid and highly specific, yielding a complex that is stable even under stringent conditions. Using the HaloLink™ Protein Array System, HaloTag® fusion proteins are expressed in a cell-free expression system and then covalently captured on hydrogel-coated glass slides derivatized with HaloTag® Ligands. The fusion proteins are captured directly from the expression reaction mixture without prior purification. Using this approach, multiple fusion proteins can be rapidly synthesized and immobilized in parallel on the slide surface, and an entire experiment including protein expression, custom array formation and protein interaction analysis can be completed in less than eight hours.

The HaloLink™ Array Six Slide System contains HaloLink™ Slides, HaloLink™ Array Gaskets and Anti-HaloTag® Antibody. To use the Six Slide System you will need to provide your own protein expression system or order the TnT® T7 Quick Coupled Transcription/Translation System (Cat.# L1170 or L1171) or TnT® SP6 High-Yield Wheat Germ Protein Expression System (Cat.# L3260 or L3261). The HaloTag® Standard Protein (Cat.# G4491) is not included with the Six Slide System but can be ordered separately.

The HaloTag® technology offers a quick and convenient way to test protein expression of HaloTag® fusion proteins by labeling with fluorescent HaloTag® TMR Ligand followed by SDS-PAGE analysis. Guidelines can be found in the HaloLink™ Protein Array Technical Manual #TM310.

The Protein G HaloTag® Fusion Protein (Cat.# G7291) is a protein consisting of recombinant Protein G from Streptococci without the albumin-binding domain and HaloTag® protein. The Protein G HaloTag® Fusion Protein, which has a molecular weight of 58kDa, can be covalently coupled to different chloroalkane surfaces, reactive and fluorescent HaloTag® ligands. This fusion protein enables an oriented attachment of antibodies to the HaloLink™ Protein Arrays.

Features - Benefits

• Fast Protein Production: Cell-free expression systems allow quick, single-tube, coupled transcription/translation for the production of the proteins of interest to be used in the protein array experiment.
• Irreversible Binding of the Captured Protein: Unlike other affinity tags, which tend to dissociate from the surface, HaloTag® fusion proteins are covalently bound to the HaloLink™ Slide.
• No Protein Purification Step: The protein of interest is immobilized directly from the cell-free expression reaction.
• Reduced Nonspecific Binding: HaloLink™ Slides have a unique hydrogel coating that is designed to prevent nonspecific binding while preserving the functionality of specifically captured proteins.
• Extensive Washing Allowed: Covalent binding of HaloTag® fusion proteins to the HaloLink™ Slide allows extensive, stringent washing that results in reduced background and a lower incidence of false positives.
• No Need for a Robotic Arrayer: The unique 50-well configuration allows multiple interactions to be studied in parallel without the need for a complex robotic arrayer.
• Highly Efficient Antibody Binding: Protein G HaloTag® Fusion Protein increases the binding of antibodies to surfaces as compared to direct capture method.

Applications

• Protein:protein interactions.
• Protein:drug interactions.
• Protein:nucleic acid interactions.
• Antibody immobilization.

Notes

Do not use the Protein G HaloTag® Fusion Protein (Cat.# G7291) in combination with the Magne™ HaloTag® Beads (Cat.# G7281); these beads are not optimized for immunoglobulin purification.

References

1. Nath, N. et al. (2008) Improving protein array performance: Focus on washing and storage conditions. J. Proteome Res. 7(10), 4475–82.
2. Los, G. et al. (2005) HaloTag® interchangeable labeling technology for cell imaging and protein capture. Cell Notes 11, 2–6.
3. Los, G. and Wood, K. (2007) The HaloTag: A novel technology for cell imaging and protein analysis. Methods. Mol. Biol. 356, 195–208.
4. Los, G. et al. (2008) HaloTag: A novel labeling technology for cell imaging and protein analysis. ACS Chem. Biol. 6, 373–82.

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